PROJECT SUMMARY The ability to genetically engineer primary T-cells creates new and highly promising prospects for tumor immunity and cancer treatment. Specifically, the transduction of T-cells with genes encoding chimeric antigen receptors (CARs) and therapeutic payloads enables T-cells to recognize antigens that are either poorly immunogenic or ignored by the immune system, and to successfully eradicate tumors. One of the major impediments of genetically engineered CAR-T-cell technology is the heterogeneity of targeted antigens and antigen escape from tumors. In this application we propose to develop and test a novel theranostic approach to augment apoptosis in tumors with heterogeneous antigen levels using antibody-based secretable payloads (ASPs) delivered by antigen-specific T cells under image guidance. We hypothesize that secretion of tumor- directed ASPs by antigen-specific CAR-T cells will induce/augment apoptosis in tumors with heterogeneous tumor antigen levels. This proposal brings together translatable antibody-based therapeutics and reporter gene imaging components in genetically engineered T-cells, allowing for non-invasive monitoring of T-cell trafficking, tumor targeting, and secretion of ASPs. We expect that genetically engineered T-cells will secrete ASPs into the tumor microenvironment, thus inducing apoptosis in a larger number of tumor targets including antigen- negative tumors. Our proposal complements the ongoing genetically engineered CAR-T cell clinical studies at MSK by exploring new strategies designed to enhance tumor targeting and T-cell effector function and to improve treatment response in tumors with heterogeneous targeted antigen levels.